CD44 is a family of cell surface proteins implicated in adhesion interactions and tumor metastasis. Multiple CD44 mRNA isoforms arise from alternative splicing of variant exons (termed v1-v10). We recently discovered a novel CD44 mRNA isoform in human papillary thyroid cancers featuring a junction between subsegments of exons 4 and 13 (v8). The sequence ACAG was repeated at both the donor and acceptor sites in the genomic DNA (G. Ermak et al., Cancer Res., 56: 1037-1042, 1996). We used reverse transcription-PCR to characterize expression of this isoform in a panel of thyroid lesions. In addition, we assayed three cryopreserved human breast cancers and two samples of normal breast tissue (from female subjects who had undergone cosmetic mammoplasty) to determine whether a similar isoform is present in breast carcinomas. Levels of the novel isoform were up-regulated in 88% of the goiters, adenomas, and papillary cancers, but were undetectable in cases of thyroiditis and absent or low-level in four samples of normal thyroid tissue. The three breast cancers each yielded a 546-bp PCR product that was not detected in normal breast tissue. The PCR product from one of the breast cancers was cloned, and sequence analysis revealed a novel mRNA isoform featuring a junction between exon 3 and an internal site within exon 13 (v8). The sequence GCTTCAG was repeated at both the donor and acceptor sites in the genomic DNA. These results show that human thyroid and breast tissues contain novel CD44 mRNA isoforms featuring unusual rearrangements at repeated sequences. Further studies are warranted to determine whether the expression of this class of isoforms correlates with growth status.